The use of recombinantly-produced therapeutic proteins has continued to increase in treating many diseases and conditions. For example, Factor VIII is a trace plasma glycoprotein that is found in mammals and is involved as a cofactor in the activation of Factor X and Factor IXa. An inherited deficiency of Factor VIII results in the bleeding disorder hemophilia A, which can be treated successfully by administration of recombinant Factor VIII.
Recombinant Factor VIII (rFVIII) can be produced by Chinese Hamster Ovary (CHO) cells transfected with a vector carrying a DNA sequence encoding the Factor VIII molecule. In some cases, rFVIII is co-produced with recombinant von Willebrand Factor (rvWF). As stated, these recombinantly-produced proteins can provide an effective treatment for hemophilia.
Conventional methods of recombinantly producing proteins involve inserting the gene responsible for the production of a particular protein of interest into host cells such as bacteria, yeast, or mammalian cells, e.g., COS or CHO cells, and then growing the cells in culture media. The cultured cells then synthesize the protein of interest. Traditional bacteria or yeast systems can be unable to produce many complex proteins in a functional form. While mammalian cells can reproduce complex proteins, they are generally difficult and expensive to grow, and often produce only mg/L quantities of protein. In addition, non-secreted proteins are relatively difficult to purify from prokaryotic or mammalian cells as they are not secreted into the culture medium. Accordingly, while recombinantly-produced therapeutic proteins can provide therapeutic benefits to a large number of diseases and conditions, the large-scale production of these proteins remains a challenge.
Regarding recombinant Factor VIII, particularly, this protein is expensive to produce due to the relatively low yields obtained in processes known in the art. The yield per cell tends to be low compared to the yield that might be obtained for other recombinant proteins. Generally, secreted FVIII is separated from source CHO cells, debris and DNA using depth filtration that employs charged CUNO filters. The charged depth filter binds to the FVIII product thereby reducing the final recovery of the protein.
Based on the foregoing, new techniques that enhance production and recovery of recombinantly-produced therapeutic proteins are needed.